what are the advantages and disadvantages of using recombinant protein and affinity chromatography for protein purification compared to gel filtration (size exclusion chromatography) and DEAE-sepharose chromatography (ion-exchange chromatography)?

During agarose gel electrophoresis, why does DNA move through the gel when electric current is applied? because DNA is negatively charged because a charged chemical from the loading buffer is bound to the DNA because DNA is positively charged because DNA absorbs electricity

Below is an EMSA showing four different reactions, A-D. In each tube there is some combination of labelled DNA probe, Protein X (the protein you are studying), and an antibody for Protein X. Identify which combination of components are found in each of the four reactions and explain how you determined that based on the molecular interactions being studied and your knowledge of gel electrophoresis. It is possible that multiple lanes have the same component(s). A B C D EMSA

Electrophoresis is an extremely useful procedure when applied to analysis of nucleic acids as it can resolve molecules of different sizes with relative ease and accuracy. Large molecules migrate more slowly than small molecules in agarose gels. However, the fact that nucleic acids of the same length may exist in a variety of conformations can often complicate the interpretation of electrophoretic separations. For instance, when a single species of a bacterial plasmid is isolated from cells, the individual plasmids may exist in three forms (depending on the genotype of their host and conditions of isolation): superhelical/supercoiled (form I), nicked/ open circle (form II), and linear (form III). Form I is compact and very tightly coiled, with both DNA strands continuous. Form II exists as a loose circle because one of the two DNA strands has been broken, thus releasing the supercoil. All three have the same mass, but each will migrate at a different rate through a gel. Based on your…

At higher amounts of protein, the Bradford assay is not linear. Consider the plot to the right: what is the maximum amount of protein a sample could contain and still fall within a standard curve? Briefly explain your reasoning.

what are two differences between agarose gel electrophoresis and SDS PAGE?

why does a higher agarose concentration render better resolution/separation of smaller DNA fragments? and what determines the distance of DNA fragments in gel electrophoresis?

Droplet based single cell DNA sequencing uses a microfluidic platform which generates droplets encapsulating individual cells. A micrograph of such a device [Zilionis et al, Nature Protocols 2016] is presented in Figure Q3. The channels are 25 μm deep. (a) (b) Oil Figure Q3. Micrograph of the microfluidic droplet device. Scale bars are 100 µm. Adapted from Zilionis et al, Nature Protocols 2016. Red arrowheads show individual cells, and black arrows indicate flow direction. (c) RT/lysis reagents Cells Barcoding hydrogel beads A company wants to explore the design of a prototype of the device made from PDMS. Describe a suitable fabrication process, explaining the rationale behind each individual step. Schematics can be drawn to illustrate your response. You may want to refer to the datasheets at the end of the script. Testing of the prototype was successful and the company wants to explore the feasibility of manufacturing the device from glass. Describe the fabrication process for this…

Why is it not necessary to dilute your protein samples with buffer in a bradford protein assay experiment?

Calculate these for 1 ug of plasmid DNA in a final total volume of 20 ul. All enzymes being used here are at a concentration of 10 U/µL. For double digests, give the appropriate volume for each enzyme and decide which buffer to use. (HINT: look up double digest conditions on the NEB website.) You should always calculate all volumes in advance to ensure the correct working concentrations and so that you can prepare the digests as efficiently as possible. It is a good idea to check off each component as it is added to the microfuge tube. Complete the table below, including the volume of DNA sample(s) you will need for 1 pg of the DNA (from two different methods in Experiment #3) based on the concentration(s) you determined from the OD260 value, which you will restriction digest with each of the restriction enzymes singly and in double digests in Part A. For example, the volume of DNA sample that contains 1 µg DNA for DNA sample from Part A, Experiment 3: OD260 = 0.024, dilution 500x…